RESUMO
Immature rat ventral prostate was used to identify a potent cell growth inhibitory factor in normal prostate. Both conditioned media and peptide extracts derived from ventral prostates inhibit the cellular growth and DNA synthesis in metastatic androgen-independent human prostate carcinoma cell line (PC3). The prostatic inhibitory factor was partially purified using an hydrophobic HPLC column (C18). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis (SDS-PAGE) of partially purified material revealed a major 25-kDa band. Immunoblot analysis showed similarity of the inhibitory factor to the transforming growth factor-beta (TGF-beta). Furthermore, the proliferation inhibiting activity of the prostatic factor was completely abolished by anti-TGF-beta antibodies. These data indicate that normal prostate produces a TGF-beta-like protein under non-physiological conditions.
Assuntos
Próstata/metabolismo , Neoplasias da Próstata/patologia , Fator de Crescimento Transformador beta/metabolismo , Animais , Divisão Celular , Células Cultivadas , Cromatografia Líquida de Alta Pressão , Meios de Cultivo Condicionados , Relação Dose-Resposta a Droga , Eletroforese em Gel de Poliacrilamida , Immunoblotting , Masculino , Peptídeos/química , Peptídeos/isolamento & purificação , Peptídeos/farmacologia , Próstata/química , Próstata/citologia , Ratos , Fator de Crescimento Transformador beta/imunologia , Fator de Crescimento Transformador beta/isolamento & purificação , Fator de Crescimento Transformador beta/farmacologia , Células Tumorais CultivadasRESUMO
The presence of thymidine phosphorylase in human healthy, adenomatous and cancerous prostate was demonstrated. The enzyme was responsible for the cleavage and synthesis of thymidine and for the transfer of deoxyribose from one deoxyribonucleoside to a pyrimidic base. The enzyme from normal and adenomatous prostate was retained on DEAE-Sephadex gel. In PC-3 cells, two enzymes with thymidine phosphorylase activity were present, one was retained on the gel, the second was excluded from it. Thymidine phosphorylase activity was higher in adenomatous and cancerous tissues that in healthy ones. In all tissues, the reactions of thymidine synthesis and of deoxyribose transfer were more important than that of thymidine cleavage.
Assuntos
Próstata/enzimologia , Hiperplasia Prostática/enzimologia , Neoplasias da Próstata/enzimologia , Timidina Fosforilase/metabolismo , Humanos , Masculino , Timidina/metabolismoRESUMO
The addition of bovine milk whey to the culture medium of human cancerous cells (MCF-7 and PC-3) results in a significant reduction of cells growth. Milk whey acts more efficiently on MCF-7 than on PC-3 growth. The inhibition could be due to a protein. Its identification is now on progress.
Assuntos
Adenocarcinoma/patologia , Neoplasias da Mama/patologia , Proteínas do Leite/farmacologia , Neoplasias da Próstata/patologia , Animais , Bovinos , Divisão Celular/efeitos dos fármacos , Humanos , Masculino , Células Tumorais Cultivadas/citologiaRESUMO
We have measured by a radioenzymatic assay the thymidine kinase in the cytosol of 182 primary infiltrating breast cancers. Maximal follow-up is 95 months. Thymidine kinase was found to be related to SBR grade, tumour size and absence of oestradiol receptors (RE). Univariate analysis has pointed out a significant linkage between overall or metastase free survival and thymidine kinase, using a cut-off level of 80 mU/mg protein which is the most discriminating value. Thymidine kinase appeared to be particularly useful in lymph-node-positive, RE-negative and grade 3 patients. Multivariate analysis of the overall survival and of the metastase free survival (Cox model) revealed that they were strongly related to thymidine kinase status.
Assuntos
Neoplasias da Mama/enzimologia , Timidina Quinase/metabolismo , Axila , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Citosol/enzimologia , Feminino , Seguimentos , Humanos , Metástase Linfática , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Receptores de Estradiol/análise , Receptores de Progesterona/análise , Análise de Sobrevida , Timidina Quinase/análiseRESUMO
Estrogens could act as effectors or inhibitors of protein synthesis in the rat uterus, depending on the doses given to animals. A single injection of estradiol-17 beta to immature female rats led to the increase in protein synthesis and in enzyme activities involved in DNA synthesis. Four injections, given once daily, resulted in the inhibition of enzyme activity and synthesis of all proteins but one. The 105 kD protein which showed a gradual increase with the duration of estrogen treatment could be responsible for the negative action of estrogens on uterine growth.
Assuntos
Estradiol/farmacologia , Útero/efeitos dos fármacos , Animais , DNA/biossíntese , DNA/metabolismo , DNA Polimerase II/metabolismo , Eletroforese em Gel de Poliacrilamida , Indução Enzimática , Feminino , Isoenzimas/antagonistas & inibidores , Isoenzimas/biossíntese , Isoenzimas/metabolismo , Biossíntese de Proteínas , Inibidores da Síntese de Proteínas/metabolismo , RNA/genética , RNA Polimerase II/metabolismo , Ratos , Ratos Endogâmicos , Timidina/metabolismo , Timidina Quinase/antagonistas & inibidores , Timidina Quinase/biossíntese , Timidina Quinase/metabolismoRESUMO
Two pyrimidine phosphorylase activities have been isolated from the cytosol of cultivated MCF-7 cells of a human breast cancer, by ion exchange chromatography. Both enzymes are responsible for the cleavage of thymidine into thymine and deoxyribose-1-phosphate, for the synthesis of thymidine and for the transfer of deoxyribose from d-uridine to thymine. These activities are likely to participate in the regulation of the pool of pyrimidine nucleosides required for DNA synthesis.
Assuntos
Neoplasias da Mama/enzimologia , Pentosiltransferases/metabolismo , Cromatografia por Troca Iônica , Citosol/enzimologia , Humanos , Pentosiltransferases/isolamento & purificação , Pirimidina Fosforilases , Ribosemonofosfatos/metabolismo , Timidina/metabolismo , Timina/metabolismo , Células Tumorais CultivadasRESUMO
The presence of thymidine phosphorylase was observed in healthy, adenomatous and tumoral prostatic cells. In healthy and adenomatous tissues the enzyme activity was recovered as a single peak after ion exchange chromatography on DEAE-Sephadex gel. On the contrary, two forms of thymidine phosphorylase were found in prostatic cancers, one of them, with high activity appeared consequently as a characteristic feature of prostatic tumoral cells.
Assuntos
Pentosiltransferases/análise , Hiperplasia Prostática/enzimologia , Neoplasias da Próstata/enzimologia , Timidina Fosforilase/análise , Cromatografia por Troca Iônica , Humanos , Masculino , Próstata/enzimologia , Timidina Fosforilase/isolamento & purificação , Timidina Fosforilase/metabolismoRESUMO
An unusual form of thymidine kinase has been found in cytosols from breast cancers. It differs from the adult and fetal isoenzymes of thymidine kinase in so far as its activity could not be isolated from that of thymidylate kinase. Following sucrose density gradients, electrofocusing, ion exchange chromatography or molecular sieving, both activities appeared as a single peak. These facts suggest that they could be attributed to a single multifunctional protein.
Assuntos
Neoplasias da Mama/enzimologia , Isoenzimas/isolamento & purificação , Timidina Quinase/isolamento & purificação , Linhagem Celular , Fenômenos Químicos , Físico-Química , Cromatografia de Afinidade , Feminino , Humanos , Isoenzimas/metabolismo , Núcleosídeo-Fosfato Quinase/metabolismo , Timidina Quinase/metabolismoRESUMO
The action of 5-androstene-3 beta,17 beta-diol (Adiol) on Thymidine Kinase (TK) was studied in the rat uterus. It appeared that Adiol, like 17 beta-estradiol (E2), could induce the synthesis of the fetal isoenzyme of TK. Its effects compared with those of E2, were delayed by 6 h and higher doses were required to reach the same levels of induction. The action of Adiol on TK was exclusively observed in tissues where E2 receptors were present. Moreover, Adiol could stimulate the activity of uterine DNA-polymerase alpha. These observations are additional evidence of the estrogen-like behaviour of Adiol. They suggest that it could participate in proliferative processes of tissues which are dependent on estrogens.
Assuntos
Androstenodiol/farmacologia , Androstenodióis/farmacologia , Estradiol/farmacologia , Timidina Quinase/biossíntese , Útero/enzimologia , Androstenodiol/metabolismo , Animais , Cicloeximida/farmacologia , Citosol/enzimologia , DNA Polimerase II/metabolismo , Dactinomicina/farmacologia , Indução Enzimática/efeitos dos fármacos , Feminino , Isoenzimas/biossíntese , Rim/enzimologia , Fígado/enzimologia , Tamanho do Órgão/efeitos dos fármacos , Ratos , Ratos Endogâmicos , Receptores de Estradiol/metabolismo , Útero/anatomia & histologiaRESUMO
Estimation of fetal Thymidine Kinase activity in primary tumors could be a useful parameter in the assessment of early relapses in patients with breast cancer. In a group of 39 women followed up for 42 months, the percentage of early relapses and their frequency was significantly higher in patients whose tumors had a high level of enzyme activity than in others.
Assuntos
Biomarcadores Tumorais/análise , Neoplasias da Mama/enzimologia , Isoenzimas/análise , Timidina Quinase/análise , Feminino , Feto/enzimologia , Humanos , Recidiva Local de Neoplasia/diagnóstico , PrognósticoRESUMO
The in vitro translation of RNAs extracted from immature female rat uteri 24 hrs. after one or four daily injections of 17 beta-oestradiol was carried out. The proteins labelled with (35S) methionine were separated by polyacrylamide gel electrophoresis and the resulting autoradiograms were scanned. The densitometric profile of uterine proteins obtained after a single injection of oestrogen showed a considerable increase in the majority of them, compared with controls. Following four injections of 17 beta-oestradiol the synthesis of all but one of the uterine proteins was observed. The Mr of the protein induced by repeated administration of oestrogens was about 105 kdaltons. This protein could possibly be involved in the inhibition of protein synthesis observed in the rat uterus after reiterated injections of 17 beta-oestradiol to female rats.
Assuntos
Estradiol/farmacologia , Biossíntese de Proteínas , Útero/metabolismo , Animais , Relação Dose-Resposta a Droga , Estradiol/administração & dosagem , Feminino , RNA/metabolismo , Ratos , Ratos Endogâmicos , Estimulação QuímicaRESUMO
Two isoenzymes of Thymidine Kinase were isolated from 90 human breast cancers by ion exchange chromatography. One was the enzyme present in adult tissues (TK-A), the second corresponded with the enzyme found in fetal tissues (TK-F). The presence of both isoenzymes was observed in the 90 tumors investigated. The activity of TK-A varied in the same range as in normal tissues, while the activity of TK-F varied to a larger extent and was higher than that of TK-A in a majority of tumors. Moreover, TK-F activity was independent of the presence or of the level of estradiol and progesterone receptors. Both isoenzymes were further characterized and compared.
Assuntos
Adenocarcinoma/enzimologia , Neoplasias da Mama/enzimologia , Feto/enzimologia , Isoenzimas/metabolismo , Timidina Quinase/metabolismo , Adulto , Feminino , Humanos , Isoenzimas/isolamento & purificação , Timidina Quinase/isolamento & purificaçãoRESUMO
Celiptium (Ce) is an antitumor drug used in the therapy of breast carcinomas, which are to a large extent dependent on estrogens. We have studied the effect of Ce on some proteins induced by estradiol (E2) in the rat uterus. It was observed that Ce administered at the same time or before E2, was able to inhibit the induction by E2 of fetal thymidine kinase (TK-F), of creatine kinase of brain-type (CK-BB) and of progesterone receptor (PR). When Ce was given after E2, its inhibitory effects were less evident. Results seemed to indicate that Ce could bind the acceptor sites for E2 receptor and thus inhibit the activity of E2-regulated genes. This assumption was corroborated by the fact that Ce did not modify the activity of enzymes not submitted to E2 regulation.
Assuntos
Alcaloides/farmacologia , Elipticinas/farmacologia , Antagonistas de Estrogênios , Timidina Quinase/metabolismo , Útero/enzimologia , Animais , Creatina Quinase/metabolismo , Estradiol/farmacologia , Feminino , Isoenzimas/metabolismo , Fosfoglicerato Quinase/metabolismo , Piruvato Quinase/metabolismo , Ratos , Receptores de Progesterona/metabolismo , Timidina Quinase/biossínteseRESUMO
In human breast cancers, assays on thymidine kinase activity revealed the synthesis of large amounts of d-TTP. This fact suggested the presence of thymidylate kinase closely associated with thymidine kinase. Results obtained with experimental tumors were quite different. These tumors appeared inadequate for the study on thymidine metabolism in mammary cancers.
Assuntos
Neoplasias da Mama/enzimologia , Isoenzimas/metabolismo , Neoplasias Mamárias Experimentais/enzimologia , Núcleosídeo-Fosfato Quinase/metabolismo , Fosfotransferases/metabolismo , Timidina Quinase/metabolismo , Animais , Feminino , Humanos , Cinética , Ratos , Ratos EndogâmicosRESUMO
Administration of phytooestrogens to immature female rats leads to a large increase in uterine thymidine kinase activity. That increase concerns to a large extent the fetal isoenzyme of thymidine kinase. These results confirm the estrogenic properties of phytoestrogens and allow to specify their physiological effects.
Assuntos
Estradiol/farmacologia , Reguladores de Crescimento de Plantas/farmacologia , Resorcinóis/farmacologia , Maturidade Sexual , Timidina Quinase/biossíntese , Útero/enzimologia , Zearalenona/farmacologia , Zeranol/farmacologia , Animais , Indução Enzimática , Feminino , Isoenzimas/biossíntese , Cinética , Ratos , Útero/efeitos dos fármacos , Útero/crescimento & desenvolvimentoRESUMO
Seventy five human breast cancers were examined in order to search for the presence of thymidine kinase of the fetal-type (TK-F). The presence of TK-F was evidenced in all tumors. Its activity varied from one to another tumor, but it was evident that the increased TK activity observed in mammary cancers could exclusively be related to high TK-F activity. Some relations between TK-F activity and the presence of estradiol and progesterone receptors (ER, PR) were obvious. The highest activities were observed in cancers with high level of ER and PR. Thymidylate kinase activity (d-TTP synthesis) varied in parallel with TK-F activity. In a general way, it was higher in ER+ PR+ than in ER+ PR- cancers.
Assuntos
Neoplasias da Mama/enzimologia , Timidina Quinase/análise , Neoplasias da Mama/classificação , Humanos , Isoenzimas/análise , Núcleosídeo-Fosfato Quinase/análise , Receptores de Estrogênio/análise , Receptores de Progesterona/análiseRESUMO
The action of estradiol-17 beta (E2) on thymidine kinase (TK) activity was studied in uteri from immature female rats. It was demonstrated that a single injection of E2 highly stimulated the enzyme activity which reached its maximum level 24 h after hormone administration. Physiological amounts of E2 were efficient and changes in TK activity were observed exclusively in uterus and liver. A single injection of Tamoxifen produced the same effect as E2 but repeated administration resulted in the complete inhibition of enzyme activity. Using antibiotics it was demonstrated that E2 induced the synthesis of new enzyme molecules rather than an increase in enzyme activity. This statement was corroborated by the fact that after hormone administration the increase in TK activity was preceded by an increase in RNA-polymerase activity and followed by that in DNA-polymerase alpha activity. Moreover, the separation of TK isoenzymes on DEAE-Sephadex and the use of d-CTP as inhibitor of the adult isozyme suggested that E2 induced the "fetal" form of the enzyme. In addition, it was demonstrated that TK activity in uteri from ovariectomized adult female rats was enhanced by E2 administration, and that the increase was due to the stimulation of the fetal isoenzyme. It was suggested that TK could be used as a marker of the action of estrogens and antiestrogens in target organs.
Assuntos
Estradiol/farmacologia , Timidina Quinase/metabolismo , Útero/enzimologia , Animais , Castração , Dactinomicina/farmacologia , Relação Dose-Resposta a Droga , Feminino , Isoenzimas/metabolismo , Ratos , Ratos Endogâmicos , Tamoxifeno/farmacologia , Fatores de TempoRESUMO
Inhibitory effects of Celiptium on the thymidine kinase synthesis induced by oestradiol-17 beta in the rat uterus. In the rat uterus, the synthesis of thymidine kinase specifically induced by oestradiol-17 beta was inhibited by Celiptium. The synthesis was totally inhibited when the drug was administered before the oestrogen and partially when it was administered after. These facts suggested that Celiptium was competitive to the acceptor sites for oestradiol-receptors and inhibited the expression of the thymidine kinase gene.
Assuntos
Alcaloides/farmacologia , Elipticinas/farmacologia , Estradiol/farmacologia , Timidina Quinase/biossíntese , Útero/enzimologia , Animais , Indução Enzimática/efeitos dos fármacos , Feminino , Ratos , Útero/efeitos dos fármacosRESUMO
Female rats were immunized with 17 beta-estradiol-6-carboxymethyloxime-bovine serum albumin. They developed antibodies to estradiol and, to a very low extent, antibodies to BSA. Anti-estradiol antibodies possessed tight specificity to estradiol-17 beta, without cross-reactivities with other estrogens. It was demonstrated that the specific estradiol binding in uterine and pituitary cytosols gradually decreased when antiserum titres increased. In uterine cytosols, the presence of progesterone receptor was studied using promegestone (R50 20) as ligand. No significant variations in promegestone binding were observed. Competition experiments however, questioned the permanence in immunized rats of the actual progesterone receptor or of a promegestone binding protein.